Proteolytic Regulation of Troponin T & Cardiac Function

描述(由申请人提供)： 肌钙蛋白复合体由肌钙蛋白C、肌钙蛋白I和肌钙蛋白T三个亚基组成，钙离子通过肌钙蛋白复合体调节心肌收缩。本研究项目探讨了一种新的心脏TnT翻译后修饰(CTnT)在心肌对能量危机适应中的作用。我们最近报道了心肌缺血-再灌注(Zhang等人，生物化学45：11681-94,2006年)和压力超负荷(冯等人，J.Physiol)中cTnT的N端限制性蛋白分解。586：3537-50,2008)。该修饰选择性地去除了cTnT的N-末端可变区，但保留了保守的中间和C-末端区域。对转基因小鼠心脏的初步研究表明，N端截断的cTnT(cTnT-ND)在肌丝中仍具有功能，并改变心肌的收缩能力。这一发现导致了一种假说，即cTnT N端可变区的蛋白水解性移除会在心肌细丝中产生一种功能状态，作为对能量危机的急性适应。为了验证这一假说，了解cTnT-ND的生理和病理生理学意义，本研究提出了三个具体的目标：目的研究选择性缺失N端可变区对cTnT功能的影响。我们将研究cTnT-ND与其他细丝调节蛋白的相互作用，以及它对肌原纤维ATPase的钙激活和心肌收缩的功能影响。目的研究N端截短型cTnT在代偿心功能和抗心肌缺血再灌注损伤中的作用。CTnT-ND对心脏功能的急性和慢性影响及其在克服能量危机中的作用将在体外工作心脏和体内进行研究。目的研究机械牵张心肌在诱导cTnT N末端限制性蛋白水解性切割中的作用，以了解这种翻译后调控的机制。 利用集成的实验系统，本研究将对TNT的结构与功能关系获得新的认识，并为未来开发治疗缺血性心脏病和心力衰竭的新方法奠定基础。

DESCRIPTION (provided by applicant): Cardiac muscle contraction is regulated by Ca2+ through the troponin complex that consists of three protein subunits: troponin C, troponin I, and troponin T (TnT). This research project investigates the role of a novel posttranslational modification of cardiac TnT (cTnT) in myocardial adaptation to energetic crisis. We recently reported a restricted N-terminal proteolysis of cTnT in myocardial ischemia-reperfusion (Zhang et al., Biochemistry 45:11681-94, 2006) and pressure overload (Feng et al., J. Physiol. 586:3537-50, 2008). This modification selectively removes the N-terminal variable region of cTnT but keeps the conserved middle and C-terminal regions intact. Initial studies in transgenic mouse hearts showed that the N-terminal truncated cTnT (cTnT-ND) remains functional in the myofilaments and alters myocardial contractility. This finding has lead to a hypothesis that the proteolytic removal of the N-terminal variable region of cTnT produces a functional state in cardiac muscle thin filaments as acute adaptation to energetic crisis. To test this hypothesis and understand the physiological and pathophysiological significance of cTnT- ND, three Specific Aims are proposed in this research project: Aim I will characterize the effects of the selective deletion of the N-terminal variable region on the function of cTnT. We will examine the interactions of cTnT-ND with other thin filament regulatory proteins and its functional impacts on the Ca2+ activation of myofibril ATPase and the contractility of cardiac muscle. Aim II will study the role of N-terminal truncated cTnT in compensating cardiac function and providing myocardial protection against ischemia-reperfusion injury. The acute and chronic effects of cTnT-ND on heart function and its role in overcoming energetic crisis will be investigated in ex vivo working hearts and in vivo. Aim III will investigate the role of mechanical stretch of the cardiac muscle in inducing the restricted proteolytic cleavage of cTnT N-terminal segment in order to understand the mechanisms that regulates this posttranslational regulation. Using integrated experimental systems, this study will gain new knowledge for the structure-function relationships of TnT and lay a foundation for future development of new treatment of ischemic heart disease and heart failure.